DETECTION OF SUBSTANCE-P IN HUMAN TEARS BY LASER-DESORPTION MASS-SPECTROMETRY AND IMMUNOASSAY

Purpose. To determine whether substance P is present in human tears. M ethods. Tear samples (1-2 mu l) were collected from one eye of each of 12 subjects. Two of the eyes had dry eye syndrome, two wore contact l enses and had dry eye syndrome, and eight were normal. Five of the eig ht normal eyes were scheduled to undergo excimer laser refractive surg ery, and tears were collected from these eyes before and after surgery . Tear samples were analyzed by laser desorption mass spectrometry. Po oled samples from one individual were subjected to enzyme-linked immun oabsorbent assay. Results. Laser desorption mass spectra of the 18 tea r samples displayed well defined peaks with mass to charge (m/z) ratio s ranging from 1343.7 to 1355.9 and/or 1356.9 to 1364.7, corresponding to an average m/z of 1349.8 +/- 1.13 for protonated substance P and 1 361.2 +/- 0.54 for oxidized substance P obtained from 14 mass spectra of standards formulated with substance P concentrations ranging from 1 0(-4) M to 10(-12) M. As confirmation, an enzyme-linked immunoabsorben t assay performed twice on pooled tears from one eye detected substanc e P in both replicates at a concentration of 125 pg/ml (9.26 x 10(-11) M). Conclusions. These findings demonstrate that substance P is a com ponent of tears obtained from normal eyes of men and women ranging in age from 26 to 60 years, from eyes fitted with contact lenses, from ey es with dry eye syndrome, and from eyes 1 and 2 days after excimer las er refractive surgery. Whether the concentration of substance P in tea rs varies with sex, age, or eye condition, the source of substance P i n tears, and its role in tears remains to be discovered.